Precipitation and counting of minute quantities of labeled nucleic acids as cetyltrimethylammonium salt

Sibatani, Atuhiro

doi:10.1016/0003-2697(70)90298-8

Cited by 77 publications

(16 citation statements)

References 5 publications

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“…It had been shown previously (Sibatani 1970) that the recovery of nucleic acids is dependent on NaCl concentration. Our results with seawater are consistent with results obtained with NaCl solutions.…”

Section: Discussionmentioning

confidence: 92%

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The measurement and distribution of dissolved nucleic acids in aquatic environments

Karl

Bailiff

1989

Limnology & Oceanography

179

157

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Nucleic acids (DNA and RNA) are ubiquitous components of the dissolved organic matter (DOM) pool of all oceanic, neritic, estuarine, and freshwater habitats studied to date. A new method for the quantitative determination ofdissolved nucleic acids (DNA and RNA) in water and scdimcnt samples was developed, evaluated, and utilized in a study of various marine and freshwater ecosystems. Under appropriate reaction conditions, dissolved DNA (D-DNA) and dissolved RNA (D-RNA) are efficiently removed from solution with the addition of cctyltrimethylammonium bromide (CTAB) and subsequent formation of insoluble CTA-nucleic acid salts. The insoluble salts are collected, by filtration, onto glass-fiber filters and analyzed for DNA and RNA with fluorometric and calorimetric procedures, respectively. The pcrformancc of this CTAB method is simple, reliable, and reproducible for measuring dissolved nucleic acids in natural aquatic environments. For the ecosystems investigated hcrcin, D-DNA and D-RNA concentrations ranged from 0.56 to 88 pg liter-' and 4.03 to 871 pg liter I; the ratio of D-RNA to D-DNA ranged from 4.1 to 11.5.In most aquatic environments, the organic carbon inventory is dominated by nonliving materials, present both in dissolved and particulate states. This nonliving, or detrital carbon is derived from nonpredatory losses of organic matter at all trophic levels (e.g. egestion, excretion, secretion, etc.) and from allochthonous sources (Wetzel et al. 1972). Detrital carbon food webs, initiated by the activities of heterotrophic bacteria (Pomeroy 1974), provide a mechanism for the utilization of the chemical energy contained in the nonliving carbon pool. The potential importance of this putative "microbial loop" carbon flux pathway has focused attention on studies of the concentration, molecular composition, and cycling rates of dissolved organic matter (DOM; Azam et al. 1983).

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Section: Discussionmentioning

confidence: 92%

“…We presently interpret this loss to be the result of extraccllular nucleases. If nucleases are suspected to be present in a given sample, one could add EDTA as recommended previously (Sibatani 1970). We have used this modification for samples collected in the eutrophic coastal area of the Antarctic peninsula (Bailiff and Karl in press).…”

Section: Discussionmentioning

confidence: 99%

The measurement and distribution of dissolved nucleic acids in aquatic environments

Karl

Bailiff

1989

Limnology & Oceanography

179

157

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“…It creates insoluble CTA-nucleic acid salts that can be easily separated from the solution by filtration or centrifugation techniques. Freezing of CTAB supplemented samples (Sibatini 1970) allows for easier separation of the DNA-CTA complexes from the liquid phase and prevents micelle or gel formation, which sometimes occurs if excess CTAB is added rapidly (Bellamy & Ralph 1968). NaCl addition (10 nmol I-') restricts the potential possibility of prec~pitation with CTAB of various acidic polysaccharides (Bellamy & Ralph 1968, Karl & Bail~ff 1989.…”

Section: Sample Filtrationmentioning

confidence: 99%

Determination of dissolved deoxyribonucleic acid concentration in lake water

Siuda¹,

Güde²

1996

Aquat. Microb. Ecol.

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In this paper we d~s c u s s the sultablllty of the cetyltrimethylammonium bromlde (CTAB) precipitation method for the determination of dissolved DNA (dDNA) In lake water. Factors affecting dDNA recovery from the precipitate and further DAPI fluorometr~c assay were also examined. The proposed method was particularly suitable for routine determination of dDNA concentration in low and moderately eutrophicated freshivater environments The detection 11m1t of the method v a n e d from 1.5 pg dDNA 1. ' for mesotrophlc waters to 2 5 pg dDNA I-' for e u t r o p h~c lakes. Preliminary observatlons on the concentration of dDNA in the surface water of 8 southern German lakes showed that dDNA concentrations were generally related to their trophic status and varied between 2.5 and 46.0 pg I-' in mesotrophlc and from 11.5 to 72 pg I-' in eutrophlc waters KEY WORDS. Dissolved D N A . Lake w a t e r . CTAB . DAPI

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“…16 The cetyltrimethylammonium bromide (CTAB)-DNA precipitation method is still in use, with more or less advanced modifications. [17][18][19][20] In previous work, precipitation maps of DNA and cationic surfactants of various chain length were introduced.…”

Section: Introductionmentioning

confidence: 99%

DNA−Cationic Surfactant Interactions Are Different for Double- and Single-Stranded DNA

et al. 2005

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The stability of DNA in solution and the phase behavior in mixtures with dodecyltrimethylammonium bromide (DTAB) were investigated. By means of circular dichroism, UV absorption, and differential scanning calorimetry, we found that for dilute solutions of DNA with no addition of salt the DNA molecules are in the single-stranded conformation, whereas the addition of a small amount of NaBr, 1 mM, is sufficient to stabilize the DNA double-helix. Furthermore, at higher DNA concentrations, native DNA becomes the most stable structure, which is due to a self-screening effect. By phase diagram determinations of the DNAsurfactant system, we found that the effect of salt on phase behavior mainly relates to a difference in interaction of the amphiphile between single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA). The difference in association between ss and dsDNA with surfactants of different chain lengths can be interpreted in terms of an interplay between hydrophobic and electrostatic interactions, the latter being influenced by polymer flexibility. In this way, a nonmonotonic variation can be rationalized. A crossing of the phase separation lines with DNA concentration can be rationalized in terms of a change in relative stability of ss and dsDNA. The fact that ssDNA phase separates earlier than dsDNA in association with DTAB, may serve as a basis for a method of easily separating dsDNA from ssDNA by the addition of surfactant; this is verified as monitored by circular dichroism measurements.

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Precipitation and counting of minute quantities of labeled nucleic acids as cetyltrimethylammonium salt

Cited by 77 publications

References 5 publications

The measurement and distribution of dissolved nucleic acids in aquatic environments

The measurement and distribution of dissolved nucleic acids in aquatic environments

Determination of dissolved deoxyribonucleic acid concentration in lake water

DNA−Cationic Surfactant Interactions Are Different for Double- and Single-Stranded DNA

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